Showing papers by "Margaret Sedgley published in 2006"

Improved methods in Agrobacterium–mediated transformation of almond using positive (mannose/pmi) or negative (kanamycin resistance) selection-based protocols

Abstract: A protocol for Agrobacterium-mediated transformation with either kanamycin or mannose selection was developed for leaf explants of the cultivar Prunus dulcis cv. Ne Plus Ultra. Regenerating shoots were selected on medium containing 15 μM kanamycin (negative selection), while in the positive selection strategy, shoots were selected on 2.5 g/l mannose supplemented with 15 g/l sucrose. Transformation efficiencies based on PCR analysis of individual putative transformed shoots from independent lines relative to the initial numbers of leaf explants tested were 5.6% for kanamycin/nptII and 6.8% for mannose/pmi selection, respectively. Southern blot analysis on six randomly chosen PCR-positive shoots confirmed the presence of the nptII transgene in each, and five randomly chosen lines identified to contain the pmi transgene by PCR showed positive hybridisation to a pmi DNA probe. The positive (mannose/pmi) and the negative (kanamycin) selection protocols used in this study have greatly improved transformation efficiency in almond, which were confirmed with PCR and Southern blot. This study also demonstrates that in almond the mannose/pmi selection protocol is appropriate and can result in higher transformation efficiencies over that of kanamycin/nptII selection protocols.

Increased Tomato Yield Through Pollination by Native Australian Amegilla chlorocyanea (Hymenoptera: Anthophoridae)

Abstract: Amegilla spp. (Hymenoptera: Anthophoridae) have been suggested as potential native Australian alternative to overseas used bumblebees (Bombus spp.) for pollination of tomato in greenhouses. In this study, we investigate the effectiveness of Amegilla chlorocyanea Cockerell as a greenhouse pollinator of tomato, Lycopersicon esculentum Mill. We show that 1) a single buzz by a female increases tomato weight by 11% compared with pollination by using an industrial pollination wand, 2) multiple buzzes increase tomato weight compared with a single buzz, and 3) unlimited flower visits lead to an increase in fruit weight of 21% compared with wand pollination. These results are comparable with those achieved by bumblebee pollination and demonstrate that A. chlorocyanea is a valid alternative to bumblebees for greenhouse tomato pollination in Australia.

Micropropagation of juvenile tissue of Eucalyptus erythronema × Eucalyptus stricklandii cv. 'urrbrae gem'

Abstract: Micropropagation via enhanced axillary shoot proliferation was investigated in the ornamental Eucalyptus cv. ‘Urrbrae Gem’ using in vitro germinated seedlings and was successfully achieved using woody plant medium (WPM) supplemented with 2.2 μM benzylaminopurine, 1.0 μM α-naphthaleneacetic acid, and 1.5 μM gibberellic acid (GA3), gelled with 5 g l−1 Phytagel®. Shoot proliferation was greater on WPM and QL media with GA3 compared to B5, AP, and TK media with or without GA3. GA3 was required for shoot elongation as the internodes were otherwise very short and unsuitable for multiplication or root initiation. Root initiation was improved using (1/2)WPM supplemented with 20 μM indole-3-butyric acid (IBA) over a 7 d pulse, followed by subculture to IBA-free medium, compared to placing shoots on low levels of IBA for 4–6 wk. Plantlets were successfully hardened off to the natural environment via a fogger at 67% relative humidity at 21°C for 3 d and continued to thrive as potted plants. This is the firs...

The use of ARMS PCR in detection and identification of xanthomonads associated with pistachio dieback in Australia

Abstract: Pistachio dieback occurs in the main pistachio growing areas of Australia. Xanthomonas strains belonging to the translucens group have been identified as the causal agent of the disease and two distinct groups, A and B, have been recognised within the pathogen population. In this study, specific primers for amplification of DNA of the pathogen were developed by sequencing the Internal Transcribed Spacer (ITS) region of rDNA from strains representing groups A and B, as well as from X. translucens isolated from wheat in Australia and one Xanthomonas translucens strain from orchard floor grasses. Primers were designed for amplification of DNA sequences specific to each group and a multiplex PCR test was developed that identified and differentiated strains of each group in a single PCR assay. To determine the specificity of the primers, PCR was carried out with DNA from 65 strains of the pistachio pathogen, 31 type and reference strains of Xanthomonas, and from 191 phytobacteria commonly found in and around pistachio orchards. In the multiplex PCR, a 331 bp fragment was amplified from all strains belonging to group A and a 120 bp fragment from all strains in group B. No PCR products were obtained from the other bacteria tested except for the type strain of X. translucens pv. cerealis, which has not been found in Australia. The assay was used to detect strains from both groups of the pathogen in pistachio plant material.

Genetic, phenotypic and pathogenic diversity among xanthomonads isolated from pistachio ( Pistacia vera ) in Australia

Abstract: Repetitive extragenic palindromic polymerase chain reaction (rep-PCR), sequencing of the 16S-23S rDNA internal transcribed spacer (ITS), biochemical and physiological tests, the Biolog microplate system, polyacrylamide gel electrophoresis (PAGE) of whole-cell proteins, and pathogenicity tests were used to characterize variability among xanthomonads isolated from pistachio trees suffering from bacterial dieback in four regions of Australia. ITS sequencing and rep-PCR revealed two distinct genotypes among the strains. The ITS sequencing suggested that the pistachio strains were closely related to Xanthomonas translucens pathovars, in particular X. translucens pv. poae. Results of physiological and biochemical tests, as well as Biolog microplate analysis and protein profiling, confirmed the existence of two groups. Furthermore, pathogenicity and host-range studies indicated that the two groups were biologically different. There was an association between the two groups and the geographical origin of the strains.

Identification of olive (Olea europaea L.) genotypes using SSR and RAPD markers

Abstract: Olive (Olea europaea L.) is one of the most ancient cultivated fruit tree species in the Mediterranean basin. It is a predominantly allogamous species showing a high degree of outcrossing which leads to considerable levels of heterozygosity and DNA polymorphism among individuals (Angiolillo et al., 1999; Rallo et al., 2000). The wide genetic patrimony and the large number of synonyms and homonyms in olive require precise methods of discrimination for cultivar identification and classification. Different techniques have been used to evaluate olive diversity. Morphological, agronomical or biochemical characterisation has been adopted for variability evaluation (Barone et al., 1994; Barranco et al., 2000; Ouazzani et al., 1993; Trujillo et al., 1995). The introduction of DNA markers provided a good discriminatory system, independent of environmental conditions. The random amplified polymorphic DNA (RAPD) technique has been applied in several studies to successfully distinguish between olive cultivars (Belaj et al., 2001; Fabbri et al., 1995; Guerin et al., 2002; Mekuria et al., 1999). Nowadays simple sequence repeat (SSR) have been proven to be very suitable markers for cultivar identification and identity typing in olive as they are transferable, highly polymorphic and co-dominant markers (Carriero et al., 2002; Cipriani et al., 2002; Rallo et al., 2000).

Effects of auxins on organogenesis and somatic embryogenesis from juvenile explants of Eucalyptus erythronema, E. stricklandii, and two inter-specific hybrids

Abstract: SummaryOrganogenesis and somatic embryogenesis were investigated in apex and cotyledon explants of Eucalyptus erythronema, E. stricklandii and their inter-specific hybrids Eucalyptus erythronema E. stricklandii cv. ‘Urrbrae Gem’ and ‘Hybrid 2.5’, following exposure to 1 mg 1–1 naphthaleneacetic acid (NAA) plus 1 mg 1–1 2,4-dichlorophenoxyacetic acid (2,4-D), or 3, 5 or 15 mg 1–1 NAA alone. Somatic embryogenesis was not observed macroscopically; however, structures characteristic of globular somatic embryos or root primordia were observed using light microscopy of apex explants of ‘Urrbrae Gem’ seedlings after 14 d on Murashige and Skoog (1962) (MS) medium supplemented with 3 mg 1–1 NAA. Root development was associated with explant vascular tissue and observed in all plant growth regulator (PGR) treatments, but was less on explants treated with 1 mg 1–1 NAA plus 1 mg 1–1 2,4-D than in all NAA-alone treatments. Shoot development was observed on apex explants after sub-culture on PGR-free medium, but was les...

Concentration and duration of ethylene treatment influences the response of banana to 1-methylcyclopropene

Abstract: We examined the effect of ethylene concentration and duration on 1-MCP efficacy with regards to shelf life and fruit quality in bananas (cv. Williams) from the middle section of bunches harvested during winter 2004 and summer 2005. Before storage, fruit was treated with ethylene (2, 20, 50 or 100 μL.L -1 ) for two consecutive days or 100 μL.L -1 for the first and 2 μL.L -1 for the second day, prior to 1-MCP (300 nL.L -1 ) exposure for 24 hrs at 22°C. To examine the effect of duration, bananas were treated with 100 μL L -1 ethylene for 30, 40 or 50 hrs prior to 1-MCP treatment (300 nL.L -1 ) at 22 °C. 1-MCP was most effective at increasing shelf life and firmness when fruit were treated with 100 μL.L -1 ethylene for the first day and 2 μL.L -1 for the second day. Interestingly, fruit harvested in winter initially treated with ethylene at the lowest concentration (2 μL.L -1 ) or exposed to the shorter duration of ethylene (30 hrs) did not ripen and remained green when treated with 1- MCP. However, winter-harvested bananas that were exposed to ethylene for 50 hrs had a longer shelf life compared to bananas treated with ethylene for 40 hrs. 1-MCP was only more effective in summer-harvested fruit when they were exposed to ethylene for 40 hrs with an increase in firmness. The discolouration index (DI) of 1- MCP treated fruit increased significantly when fruit were exposed to the ethylene for shorter durations than 50 hrs in winter, but no differences were observed in DI of 1-MCP treated bananas that were ripened with ethylene at different concentrations. These observations suggest that the efficacy of 1-MCP to improve shelf life and quality of bananas is reliant on not only the harvest season of fruit but also the concentration and duration of ethylene application prior to 1-MCP usage.